This application describes the first large-scale analysis of the genetic requirements for establishment of a human symbiont within a mammalian host. Microbes constitute -90 percent of the cells in the human body. The majority live in the gastrointestinal tract, where they play critical roles in gut development, immune maturation, and energy harvest. The composition of the mammalian gut microbiota is dynamically linked to the development of obesity, but little is known about the mechanisms that control colonization and establishment of human gut symbionts under normal or pathological conditions. The experiments proposed here test the hypothesis that human symbionts possess dedicated mechanisms that allow colonization despite continuous perfusion, intense resource competition, and immune surveillance. Applying a novel transposon mutagenesis system (Aim 1a) to a prominent human gut symbiont and monitoring the relative fitness of each member of this mutagenized population in germfree mice (Aim 1b) will identify these genetic determinants. Manipulation of microbial community complexity and host physiologic status (through genetic and diet-induced obesity, Aim 2) will connect the obesity-dependent restructuring of the gut microbiota with the mechanisms that determine fitness for symbionts in the obese or lean gut. Identification of these mechanisms will help establish strategies to optimize the contribution of the gastrointestinal microbiota to human health. PUBLIC HEALTH RELEVANCE: The microbial community that resides in the gastrointestinal tract of every human mediates a significant proportion of our caloric intake, and the composition of this community is dynamically linked to the development of obesity. This application uses genetic and diet-induced mouse models of obesity to identify and characterize the adaptive mechanisms employed by human microbial symbionts in the mammalian gut. Identifying these genetic determinants of symbiosis in single- and multi-species gut microbial communities will facilitate efforts to manipulate these communities in ways that increase human health.